“You can take a spoonful of that protein and it
generates as much torque as a Mercedes engine.”

Suspend your disbelief. The protein adenosine triphosphate synthase,
better known as ATPase,
is nature’s smallest rotary motor.
“You can take a spoonful of that protein,” says
biophysicist Klaus
Schulten of the University of Illinois Urbana-Champaign,
“and it generates as much torque as a Mercedes engine.”

A remarkable molecular motor that in the laboratory produces torque
from chemical fuel, ATPase works the other direction in humans —
converting torque into ATP,
the basic fuel of life, the chemical energy that fuels muscle
contraction, transmission of nerve messages and many other functions.
Probably the most abundant protein in all living organisms, ATPase is the power
plant of metabolism. In an active day, an adult human can produce and
consume its body weight or more of
ATP, nearly all of it produced by ATPase.

The 1997
Nobel Prize in Chemistry recognized Paul Boyer and John Walker for
their work in assembling, for the first time, a detailed picture of
ATPase and how
this tiny molecular machine does its job. Subsequent research has
added to the picture, but many challenging questions remain.

Imagine we’re from Mars, says Schulten, director of the Theoretical and Computational
Biophysics Group at the University of Illinois Beckman Institute,
and we want to understand how a car engine works, but the engine is
infinitesimally small. “It’s nearly impossible to see the
details in motion. The only way is to use the computer to simulate it,
and then we can recognize the combustion process driving the car
engine. It appears that
ATPase is kind of a combustion engine too.”

Klaus Schulten (left) and Markus Dittrich, University of Illinois, Urbana-Champaign.

To see what they could see, Schulten and graduate student Markus Dittrich turned to
Jonas, PSC’s
128-processor HP Marvel system, which is dedicated to
biomedical research. With the exceptional capability of this system,
they were able to simulate “combustion” in ATPase with quantum
theory — to get a picture of how electrons move from
atom-to-atom during chemical reactions.

“If you want to do careful calculations,” says
Schulten, “you have to invest a lot of computing power.
It would make no sense to approach this important problem with
cheap methodology. We decided to do the most advanced
calculation that people do today when simulating a biological
reaction.” Jonas’s very fast EV7 processors and
large shared memory made the quantum calculations feasible, and
the resulting study turned up invaluable new information about
nature’s tiniest motor.

The Wheel Spins Round and Round

This cartoon schematic shows the F0 part of ATPase (blue)
sitting in a membrane, where proton (H+) concentration
outside (below) the membrane greater than on the inside causes
F0 to rotate. The axle (red) of F1 also rotates and
triggers rotary "combustion" in three active sites,
contained within alternating subunits (green) of F1's
hexagonal active-site complex.

Like most motors, ATPase has moving and non-moving parts.
There’s a wheel that spins, similar to a millwheel, to
turn an axle that revolves inside a hexagonal cluster, in which
there are three combustion chambers (active sites), each of
which, in sequence, charges up with chemical raw materials
— adenosine diphosphate (ADP) and phosphate — and
“fires” to produce ATP.

The wheel part of the protein is called F0. In humans and other
mammals, F0 resides in the membrane of mitochondria,
microscopic structures inside the cell. In bacteria, where
ATPase works reversibly both as an ATP generator and an
ATP-fueled motor, F0 sits in the cellular wall. In both cases,
it forms a channel for protons to flow through the wall, a flow
which — much like water turning a millwheel —
causes F0 to rotate.

The other main part of ATPase’s structure, called F1,
extends into the cellular or mitochondrial interior. F1
includes a central stalk — the axle — that is
coupled to and turns with F0. The other end of this axle
revolves inside F1’s hexagonal cluster, which contains the
three active sites where “combustion” occurs.

“F0 spins, and this spins the F1 central stalk,” says
Dittrich, “and this leads to ATP synthesis.” The axle
spins in one direction (clockwise when viewed from the membrane side)
during ATP production. What’s not only fascinating — but
also a large benefit for understanding the protein — is that the
reverse reaction also works in the laboratory. Put F1, a very large
protein by itself, in solution with ATP, and ATP will
“hydrolyze” inside F1’s active sites into ADP and
phosphate and the axle will spin counterclockwise.

THIS AMINO ACID — THE ARGININE FINGER — SEEMS TO OPERATE
LIKE A SPARK PLUG

Because ATP hydrolysis is a chemical mirror of synthesis and more
amenable to laboratory study, it offers an invaluable back-door
approach to gaining knowledge about ATPase, and this reaction is what
Schulten and Dittrich set out to simulate.

Zooming-In on the Spark Plug

One of the intriguing questions about F1-ATPase is how its three
combustion chambers cooperate with each other. During each 360-degree
rotation of the F1 axle, each active site, one after another, changes
structure and thereby alters its function. For the hydrolysis
reaction, each site is open to bind with an
ATP molecule, closes to
hold it during breakdown into constituent products, and opens again
during product release. With each rotation, three ATP molecules
hydrolyze, one at a time. Schulten and Dittrich hoped to shed light on
the atom-by-atom details of this rotary process.

Another challenging question concerns how the hydrolysis reaction
causes the F1 axle to spin. “We don’t understand how the
chemical reaction is coupled to rotation of the axle,” says
Schulten. “That’s still a big mystery.”

To get at the crucial details of bonds breaking and reforming during a
chemical reaction requires quantum theory. Schulten and Dittrich
therefore used a method called
QM/MM (quantum mechanics/molecular mechanics), which made it
possible to simulate the entire subunit of F1 that houses the active
site, but used quantum theory selectively like a zoom lens to zoom-in
on the active site itself, where “combustion” occurs.

In this representation of the molecular structure of
F1-ATPase, the rotating stalk (red) protrudes from a
hexagonal cluster composed of alternating alpha (yellow) and
beta (green) subunits, three of each, within which the stalk
rotates. The three active sites are in the beta subunits.

The active-site subsystem for the QM part of the simulations
(blue) includes water molecules. The blowup represents this
active-site subsystem with ATP.

In total, they simulated two different configurations of the F1-ATPase
active-site subunits, more than 8,000 atoms each, while the QM
calculations zoomed in on the combustion chamber. Employing up to 32
of Jonas’s processors at a time, with an extensive series of
simulations, they used over 12,000 hours of computing time. The
outcome is several key findings on a crucial biological system.

ATP in solution without ATPase is extremely slow to hydrolyze, taking
as long as a week. With ATPase, this reaction goes about 100 billion
times faster, a huge speedup that researchers have been hard pressed
to explain. The Jonas simulations reinforce Schulten and
Dittrich’s finding (in a prior simulation study) that ATPase
hydrolysis — in which water attacks ATP to break it down —
is carried out by two water molecules in concert, rather than only
one, as had been thought. This finding — unobservable in
laboratory experiment — is a big step toward explaining
ATPase’s remarkable catalytic efficiency.

“In order to have this kind of concerted action of two water
molecules,” explains Dittrich, “they have to be arranged
in a particular way. This is accomplished by ATPase and won’t
happen in solution because there it’s unlikely the water
molecules will assume this special conformation. If we didn’t
have this mechanism, the reaction would take place on a much slower
time-scale and wouldn’t lead to the observed physiological
rates.”

The simulations also show, unexpectedly, that there’s no energy
change in the active site as ATP breaks down into its reaction
products — a finding that goes further than experiments in
establishing that the reaction itself doesn’t provide any force
toward making the F1 axle rotate. “It’s not the chemical
event that drives the rotation,” says Dittrich, “which
means we have to look at other possibilities.” The two remaining
possibilities are when ATP binds to the active site or when the
reaction products are released.

Perhaps most importantly, the simulations reveal that one of the amino
acids in the active site of F1-ATPase — arginine — appears
to play a key role in coordinating the timing among the three active
sites. This amino acid — referred to as the arginine finger
— seems to operate like a spark plug. It shifts its position
depending on whether ATP or the reaction products are in the
combustion chamber. “We think that the findings,” says
Schulten, “particularly with respect to the arginine finger,
could well prove to be a crucial part of this puzzle.”